The goal of this project is to evaluate strategies to eliminate HIV and latent reservoirs in a monkey model of HIV/SHIV infection. Recent promising vaccine trials have failed to protect from HIV, emphasizing the importance of developing alternative therapies. One such alternative therapy is the genetic modification of hematopoietic cells to make them resistant to HIV infection. Genetic modification of hematopoietic stem cells (HSCs) with zinc finger nucleases targeting the CCR5 gene loci have been shown to reduce the severity of disease in an HIV mouse model system. Thus, we propose a study aimed at determining the potential clinical application of a gene modifying approach targeting CCR5 as an antiviral therapy with the overall goal being the eradication of the viral reservoir in HIV[+] patients. We will study the effect of CCR5[-/-] repopulating cells on the latent reservoir using a clinically relevant nonhuman primate AIDS model, the pigtailed macaque (M. nemestrina). This model will allow us to carefully and thoroughly analyze the impact of CCR5[-/-] hematopoietic reconstitution on the control of HIV/SHIV. Here we will address several crucial questions regarding the feasibility and translatability of this approach. We will evaluate methods to efficiently modify HSCs in a large animal model, determine the engraftment potential of ZFN-modified HSCs and the level of ZFN-modified repopulating cells necessary for efficient control of HIV. Given the high clinical relevance of the SHIV macaque model, these studies should be readily translatable to the understanding of latency in HIV/AIDS patients. This project complements and interacts closely with the other projects and cores. Specifically, data from Project 1 will serve as a baseline for the latent reservoir in monkeys, Project 2 will provide and develop novel reagents. Project 4 will develop anti HIV strategies that can be incorporated into the monkey studies and Project 5 will develop novel delivery approaches which can also be tested in monkeys. This project will make use of all the cores. We propose the following four aims: 1) Determine the optimal approach for genetically modifying hematopoietic stem cells with zinc finger nucleases. 2) Determine the engraftment potential of ZFN-modified CCR5[-/-] repopulating cells in macaques. 3) Establish means to efficiently and safely increase the percentage of CCR5-modified cells. 4) Determine if transplantation with CCR5[-/-] CD34[+] cells can prevent infection and provide long-term control.